By the methods of X-ray protein crystallography, we propose to obtain the structure-function relationship in terms of the refined three-dimensional structure of penicillo-pepsin (an extracellular acid protease from Penicillium zanthinellum) and structural details of the way in which penicillopepsin interacts with its substrates, activators, and inhibitors. We also plan to establish the evolutionary homology as expressed in the secondary and tertiary structures among various acid proteases of fungal and mammalian origins. Penicillopepsin is one of yet a small number of well-characterized acid proteases, the other representative being notably the mammalian pepsins and chymosin. As interpreted from the 2.8 angstrom map, penicillopepsin comprises 323 amino acid residues in a single polypeptide chain which folds into two distinct lobes separated by a 30-angstrom units long cleft which is believed to be the extended binding site. The catalytic residues Asp-32 and Asp-215 are located in this cleft and their carboxyl groups in close contact through a hydrogen bonding. We will attempt to introduce substrates, inhibitors, and activators into the enzyme by soaking or co-crystallization method and to locate these compounds with the aid of difference Fourier technique. Chemical modification and pH dependence studies should reveal the structural variations. We will also try to recrystallize penicillopepsin at low ionic strength and determine its structure by molecular replacement methods. The conformational differences, if any, in the active center will be compared for crystals grown from both low and high salt concentrations. The results derived from the proposed work will be of great use in elucidating the principles which govern the enzymatic action of acid proteases.